Classifications

• • B—PERFORMING OPERATIONS; TRANSPORTING
  • B03—SEPARATION OF SOLID MATERIALS USING LIQUIDS OR USING PNEUMATIC TABLES OR JIGS; MAGNETIC OR ELECTROSTATIC SEPARATION OF SOLID MATERIALS FROM SOLID MATERIALS OR FLUIDS; SEPARATION BY HIGH-VOLTAGE ELECTRIC FIELDS
  • B03C—MAGNETIC OR ELECTROSTATIC SEPARATION OF SOLID MATERIALS FROM SOLID MATERIALS OR FLUIDS; SEPARATION BY HIGH-VOLTAGE ELECTRIC FIELDS
  • B03C1/00—Magnetic separation
  • B03C1/005—Pretreatment specially adapted for magnetic separation
  • B03C1/01—Pretreatment specially adapted for magnetic separation by addition of magnetic adjuvants
• • B—PERFORMING OPERATIONS; TRANSPORTING
  • B82—NANOTECHNOLOGY
  • B82Y—SPECIFIC USES OR APPLICATIONS OF NANOSTRUCTURES; MEASUREMENT OR ANALYSIS OF NANOSTRUCTURES; MANUFACTURE OR TREATMENT OF NANOSTRUCTURES
  • B82Y25/00—Nanomagnetism, e.g. magnetoimpedance, anisotropic magnetoresistance, giant magnetoresistance or tunneling magnetoresistance
• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
  • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
  • C12N15/09—Recombinant DNA-technology
  • C12N15/10—Processes for the isolation, preparation or purification of DNA or RNA
  • C12N15/1003—Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor
  • C12N15/1006—Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor by means of a solid support carrier, e.g. particles, polymers
• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
  • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
  • C12N15/09—Recombinant DNA-technology
  • C12N15/10—Processes for the isolation, preparation or purification of DNA or RNA
  • C12N15/1003—Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor
  • C12N15/1006—Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor by means of a solid support carrier, e.g. particles, polymers
  • C12N15/1013—Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor by means of a solid support carrier, e.g. particles, polymers by using magnetic beads
• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
  • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
  • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
  • C12Q1/6806—Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay
• • H—ELECTRICITY
  • H01—ELECTRIC ELEMENTS
  • H01F—MAGNETS; INDUCTANCES; TRANSFORMERS; SELECTION OF MATERIALS FOR THEIR MAGNETIC PROPERTIES
  • H01F1/00—Magnets or magnetic bodies characterised by the magnetic materials therefor; Selection of materials for their magnetic properties
  • H01F1/01—Magnets or magnetic bodies characterised by the magnetic materials therefor; Selection of materials for their magnetic properties of inorganic materials
  • H01F1/03—Magnets or magnetic bodies characterised by the magnetic materials therefor; Selection of materials for their magnetic properties of inorganic materials characterised by their coercivity
  • H01F1/032—Magnets or magnetic bodies characterised by the magnetic materials therefor; Selection of materials for their magnetic properties of inorganic materials characterised by their coercivity of hard-magnetic materials
  • H01F1/10—Magnets or magnetic bodies characterised by the magnetic materials therefor; Selection of materials for their magnetic properties of inorganic materials characterised by their coercivity of hard-magnetic materials non-metallic substances, e.g. ferrites, e.g. [(Ba,Sr)O(Fe2O3)6] ferrites with hexagonal structure
  • H01F1/11—Magnets or magnetic bodies characterised by the magnetic materials therefor; Selection of materials for their magnetic properties of inorganic materials characterised by their coercivity of hard-magnetic materials non-metallic substances, e.g. ferrites, e.g. [(Ba,Sr)O(Fe2O3)6] ferrites with hexagonal structure in the form of particles
• • H—ELECTRICITY
  • H01—ELECTRIC ELEMENTS
  • H01F—MAGNETS; INDUCTANCES; TRANSFORMERS; SELECTION OF MATERIALS FOR THEIR MAGNETIC PROPERTIES
  • H01F1/00—Magnets or magnetic bodies characterised by the magnetic materials therefor; Selection of materials for their magnetic properties
  • H01F1/01—Magnets or magnetic bodies characterised by the magnetic materials therefor; Selection of materials for their magnetic properties of inorganic materials
  • H01F1/03—Magnets or magnetic bodies characterised by the magnetic materials therefor; Selection of materials for their magnetic properties of inorganic materials characterised by their coercivity
  • H01F1/032—Magnets or magnetic bodies characterised by the magnetic materials therefor; Selection of materials for their magnetic properties of inorganic materials characterised by their coercivity of hard-magnetic materials
  • H01F1/10—Magnets or magnetic bodies characterised by the magnetic materials therefor; Selection of materials for their magnetic properties of inorganic materials characterised by their coercivity of hard-magnetic materials non-metallic substances, e.g. ferrites, e.g. [(Ba,Sr)O(Fe2O3)6] ferrites with hexagonal structure
  • H01F1/11—Magnets or magnetic bodies characterised by the magnetic materials therefor; Selection of materials for their magnetic properties of inorganic materials characterised by their coercivity of hard-magnetic materials non-metallic substances, e.g. ferrites, e.g. [(Ba,Sr)O(Fe2O3)6] ferrites with hexagonal structure in the form of particles
  • H01F1/112—Magnets or magnetic bodies characterised by the magnetic materials therefor; Selection of materials for their magnetic properties of inorganic materials characterised by their coercivity of hard-magnetic materials non-metallic substances, e.g. ferrites, e.g. [(Ba,Sr)O(Fe2O3)6] ferrites with hexagonal structure in the form of particles with a skin
• • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
  • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
  • Y10T—TECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION
  • Y10T428/00—Stock material or miscellaneous articles
  • Y10T428/29—Coated or structually defined flake, particle, cell, strand, strand portion, rod, filament, macroscopic fiber or mass thereof
  • Y10T428/2982—Particulate matter [e.g., sphere, flake, etc.]
  • Y10T428/2989—Microcapsule with solid core [includes liposome]
• • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
  • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
  • Y10T—TECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION
  • Y10T428/00—Stock material or miscellaneous articles
  • Y10T428/29—Coated or structually defined flake, particle, cell, strand, strand portion, rod, filament, macroscopic fiber or mass thereof
  • Y10T428/2982—Particulate matter [e.g., sphere, flake, etc.]
  • Y10T428/2991—Coated
  • Y10T428/2993—Silicic or refractory material containing [e.g., tungsten oxide, glass, cement, etc.]
• • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
  • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
  • Y10T—TECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION
  • Y10T428/00—Stock material or miscellaneous articles
  • Y10T428/29—Coated or structually defined flake, particle, cell, strand, strand portion, rod, filament, macroscopic fiber or mass thereof
  • Y10T428/2982—Particulate matter [e.g., sphere, flake, etc.]
  • Y10T428/2991—Coated
  • Y10T428/2993—Silicic or refractory material containing [e.g., tungsten oxide, glass, cement, etc.]
  • Y10T428/2996—Glass particles or spheres

Definitions

• the subject of the application is a preparation of particles with a glass surface, a ner method for producing such a preparation and a ner method for cleaning nucleic acids with the aid of this preparation.
• nucleic acids have increasingly come into the focus of interest in medical diagnostics.
• detection methods have been developed in which the presence or absence of certain nucleic acids is assessed as a sign of a disease.
• these include e.g. B. Evidence of infectious organisms, e.g. B. of viruses or bacteria in body fluids, but also the detection of mutations in genomic nucleic acids, eg. B. in oncology.
• nucleic acids are present in the sample material usually used in very low concentrations. For this reason, different methods for isolating the nucleic acids from other sample components, such as proteins or other cellular components, which partially interfere with the subsequent detection methods, have been developed.
• WO 91/12079 describes a method for isolating nucleic acids with the aid of magnetic particles made of cellulose and iron oxide, the particle size being between 1 and 10 ⁇ m. These particles do not contain a glass surface and are only suitable for isolation with the precipitation of nucleic acids. However, aggregation includes a large number of sample components that interfere with later process steps.
• EP-B-0389 063 proposes a method in which the sample is mixed with a mixture of a chaotropic guanidinium salt and silica particles. Under these conditions, nucleic acids bind to the silica surface relatively independently of the sequence. The remaining sample components can be washed off and the nucleic acids can then be eluted.
• WO 96/41811 describes magnetic particles with an essentially pore-free glass surface for the sequence-independent purification of nucleic acids.
• the particles used there have a core, preferably with magnetite as the magnetic material, and have a preferred grain size of between 10 and 60 ⁇ m.
• Magnetite in crystals larger than approx. 30 to 50 nm shows hard magnetic properties. Permanent magnetism is induced by an external magnetic field. Particles with such hard magnetic cores show the properties of a small permanent magnet after the first exposure to an external magnetic field. In suspension, such particles attract each other and form larger units. Under the influence of an external gravitational field, these larger units sediment faster than the particles as individuals. This is disadvantageous since longer incubations require frequent redispersion.
• WO 96/41840 describes pigments that have a glass surface with a thickness of at least 0.8 ⁇ m. Zinc compounds are also proposed as a glass-forming component. This produces pigment particles with a particle size of preferably 2 to 20 ⁇ m.
• nucleic acid purification or / and to further increase the yield in nucleic acid purification, or / and to provide particles for nucleic acid purification which, even after the action of an external magnetic field, show only a very slight tendency to accumulate and sediment in the gravitational field just as slowly as those which have never been exposed to a magnetic field were.
• the invention relates to a preparation comprising particles with a glass surface, more than 75% by weight of these particles having a grain size of between 0.5 and 15 ⁇ m.
• the invention further relates to a method for producing a preparation of particles comprising a core coated with a gel layer or a glass layer and a method for purifying nucleic acids with the aid of the preparation according to the invention.
• Another object of the invention is a method for producing and preparing particles with a superparamagnetic core.
• Another object of the invention is a method for producing and preparing particles with a magnetic, preferably a soft magnetic metallic core.
• the person skilled in the art refers to solid materials with a small diameter. These particles preferably have an essentially spherical surface. Here, platelet-shaped and thread-like particles in the dimensions given below are also to be understood as particles.
• a core pigment portion
• Such cores preferably contain metal oxides such as aluminum oxide, iron oxide, chromium oxide, copper oxide, manganese oxide, lead oxide, tin oxide, titanium oxide, zinc oxide and zirconium oxide or metals such as Fe, Cr, Ni or magnetic alloys.
• composition of this core is less important for the function of the particles according to the invention, since the core is coated with a glass surface, so that the core does not come into direct contact with the sample from which the nucleic acid is to be isolated.
• Such kernels are commercially available. If the core contains Fe 3 O 4 (magnetite) or Fe 2 O 3 (maghemite) or Fe or Cr or Ni or magnetic alloys, these cores are magnetic.
• Suitable materials that are referred to as soft magnetic are metals based on the pure elements Fe, Ni, Cr and alloys thereof, preferably based on Ni. Examples of such alloys are known under the name Permalloy. They consist of 70 to 80% Ni with additions of Cr, Cu and Mo. Particles made of magnetically soft material do not attract one another or only negligibly in the absence of an external magnetic field.
• Fine-particle metal powders are very reactive. There is a risk of spontaneous combustion in air, they are considered pyrophoric. It was therefore very surprising that such finely divided metal particles could be coated with a glass layer by the sol-gel process without the magnetic properties changing significantly.
• Carbonyl iron powders are particularly preferably used as the metal powder, with types reduced in H 2 having magnetically particularly preferred properties. Caronyl iron whiskers also have preferred properties.
• Metal powders preferably have a grain size of between 10 nm and 100 ⁇ m, particularly preferably of between 200 nm and 8 ⁇ m.
• a glass surface in the sense of the present invention consists of a silicon-containing amorphous material.
• the glass preferably contains one or more of the following components (in mol%):
• B 2 O 3 (0-30%), Al 2 O 3 (0-20%), CaO (0-20%), BaO (0-10%), K 2 O (0-20%), Na. 0 (0-20%), MgO (0-18%), Pb 2 O 3 (0-15%), ZnO (0-6%).
• Borosilicate glasses which are particularly preferred from the point of view of the yield of nucleic acids have a zinc oxide content of 2-6, preferably of about 4 mol%.
• the glass layer particularly preferably consists of 68-79 mol% SoO 2 , 15- 5 mol% B, O 3 , 6-2.5 mol% total amount of K 2 O and Na.0, 4-1 mol% CaO, 8-2 mol% Al 2 O 3 , 6-2 mol% ZnO. Glasses which are formed by the so-called gel-sol process and subsequent drying and compacting of the layer formed are particularly preferred in the sense of the invention. The basic features of this process are known and have been described in e.g. B. in CJ Blinker, GW Scherer "Sol Gel science - The physics and chemistry of Sol Gel Processing", Academic Press Inc. 1990 and Sol-Gel Optics, Processing and Applications Lisa C. Klein Ed.
• alkoxides of network-forming components e.g. B. SiO 2 , B 2 O 3 , Al 2 O 3 , TiO 2 , ZrO 2 and ZnO together with oxides and salts of other components, e.g. B. in alcoholic solution, submitted and hydrolyzed.
• the hydrolysis process of the starting components is started by adding water.
• the reaction proceeds relatively quickly, since the alkali ions have a catalytic effect on the rate of hydrolysis of the silicic acid ester.
• the resulting gel can be dried and compressed into a glass by a thermal process.
• the quantitative ratio of sol / pigment has a considerable influence on the yield of magnetic pigment according to the invention. There are limits to the fact that the pigment content is so low that a mass that can still be pumped and sprayed is formed. If the pigment content is too low, the fine content, e.g. B. of non-magnetic material too large and disturbing. 10 to 45 g pigment / 100 ml sol were found to be appropriate quantitative ratios with regard to the pigment yield.
• the slurry is preferably sprayed through a nozzle and the aerosol is dried on a falling section.
• the nozzle is preferably heated to accelerate drying of the slurry.
• the nozzle temperature is preferably approximately 120 to 250 ° C. A compromise is found by sufficient evaporation speed, but avoiding splashing.
• the compression temperature should be as high as possible. However, if it is too high, the particles stick together and agglomerates form which should be sieved out.
• the addition of zinc in the layer surprisingly increases the melting point, so that a higher compression temperature (between 710 and 800 ° C) is possible.
• the post-treatment in air leads to a loss of magnetic properties at too high temperatures, which is why excessive temperatures should be avoided.
• other temperatures are possible when zinc is added (preferably between 150 and 250 ° C).
• magnetic cores which are substantially smaller can also be used as magnetic cores in the processes described in WO 96/41811.
• nanoscale cores e.g. Magnetite
• the lower limit of the core size results from the manageability of the cores, especially their tendency to form aggregates.
• the nuclei are preferably larger than 5 nm, particularly preferably 7 nm.
• the magnetic behavior of the nanoscale nuclei is referred to as superparamagnetic. The particles obtained sediment rapidly under the influence of an external magnetic field.
• the sedimentation speed in the gravitational field does not differ from the sedimentation speed in the gravitational field before exposure to the external magnetic field.
• the advantage here is that longer incubation times in suspension are possible without having to be mixed and resuspended again.
• a preparation of core particles in which more than 75% by weight of the core particles have a particle size of between somewhat less than 0.5 and somewhat less than 15 ⁇ m is used in the sol / gel process.
• the core particles must be as much smaller than the glass-coated particles as the thickness of the glass layer is.
• the glass layer will be between 5 nm and 1 ⁇ m thick, depending on the circumstances chosen, such as the ratio of gel to core particles. On average, the glass layer should be between 0.2 and 0.3 ⁇ m thick.
• a preparation containing particles with a glass surface is particularly preferred, with more than 75% by weight of these particles having a grain size of between 2 and 15 ⁇ m.
• the proportion of particles with the determined grain size is particularly preferably greater than 90% by weight.
• Magnetic core particles are particularly preferably used.
• the preparation according to the invention has the advantage that preferably more than 95% by weight of the particles with a grain size of between 0.5 and 15 ⁇ m, preferably between 2 and 15 ⁇ m, are magnetic. This means that the proportion of non-nuclear particles is drastically reduced compared to the known processes. This can be seen from the fact that there are only a few non-magnetic particles. This means that it is practically no longer necessary to separate the non-magnetic particles formed from the magnetic particles before the preparation is used in processes for the purification of nucleic acids. This means a simplification in the manufacturing process.
• the preparation according to the invention can be characterized in that preferably less than 50% of the particles have a grain size of less than 2 ⁇ m.
• the non-magnetic fine fraction which makes up a large relative fraction for small grain sizes, is greatly reduced.
• less than 2% of the particles have a grain size of less than 0.5 ⁇ m.
• the preparation according to the invention may also contain further non-glass-containing constituents, such as, for. B. buffer substances or a suspending agent, e.g. B. water or alcoholic solutions of water.
• the glass layer of the particles of the preparation according to the invention preferably contains a proportion of between 2 and 6 mol%, particularly preferably 4 mol%, of zinc oxide. This can be achieved in that the proportion of zinc oxide in the solid mass of the sol is in this order of magnitude compared to the proportions of the other solid components.
• the proportion of zinc oxide increases with a reduction in the proportion of Boron oxide, especially after prolonged heating, since boron oxide is already volatile under the manufacturing conditions.
• Particles that have a glass layer in which the proportion of zinc oxide is between 2 and 6 mol% have proven to be particularly effective in the purification of nucleic acids.
• the yield of nucleic acids could be partially increased by 50% compared to the same glass layer without zinc oxide.
• the invention also relates to a method for producing a preparation of particles with a core coated with a gel layer containing less than 5% by weight of coreless particles, comprising the steps of suspending core particles in a sol using a core particle preparation and spray-drying the suspension with gel formation, wherein the core particle preparation consists of 75% by weight of particles with a grain size of between 0.5 and 15 ⁇ m, preferably between 2 and 15 ⁇ m.
• a method has proven to be particularly advantageous in which initially a sol of tetraalkyl orthosilicates, alkyl borates. Aluminum alcoholates and alkali alcoholates are made in ethanol and this mixture is heated with calcium. The mixture is then hydrolyzed by adding water. The core particles are added in solid form to the sol formed in this way and suspended, preferably with ultrasound.
• the suspension is then gel-formed in a spray-drying process in which the nozzle is heated and in which essentially particles are formed which contain 1 to only a few core particles per particle (preferably less than 1% of the particles contain more than 10 Core particles), sprayed.
• the spray product is then heated to compress the gel into a glass.
• the addition of zinc oxide to the gel has a considerable advantage.
• the densification can be carried out at higher temperatures than those without the addition of zinc, since the softening point of the resulting glass is higher. This makes it easier to drive out organic residual components from the materials used.
• the invention also relates to a method for the purification of nucleic acids by non-covalent binding of the nucleic acids from a sample to particles with a glass surface, removal of unbound sample components and elution of the bound nucleic acids from the glass surface, a preparation according to the invention being used.
• the process becomes particularly simple if the particles are magnetic.
• WO 96/41811 Methods for purifying nucleic acids using magnetic particles with a glass surface are described in WO 96/41811. This disclosure is fully referred to here.
• Clinical samples such as blood, serum, mouthwash, urine, cerebral fluid, sputum, stool, plasma, punctures or bone marrow samples are particularly suitable as samples for the cleaning method according to the invention.
• the preferred sample material is serum.
• the sample if necessary after lysis of any cellular structures it contains and digestion of disturbing sample components, is mixed with the preparation according to the invention, e.g. B. in the form of a certain amount of a suspension of the particles.
• the liquid is removed together with the unbound sample components and, if desired, the particles are washed to remove residues.
• the nucleic acids still bound to it are removed from the surface by elution with a liquid in which the nucleic acids dissolve well.
• the resulting liquid can now be processed as desired, in particular used in amplification processes, such as. B. PCR, since most enzyme inhibitors were removed during the purification process.
• the removal of the liquid from particles with the nucleic acids is particularly easy since the particles are collected with the aid of a magnet and can be held while the liquid is removed. If the particles are not magnetic, they can be separated from the liquid by filtration with a suitable filter.
• the mixture is then heated with stirring until strong reflux.
• a mixture of a total of 583 ml of ethanol and 233 ml of water is added dropwise over 30 minutes.
• the sol is poured into an open container and 1200 g of pigment IRIODIN 600 Black Mica (manufacturer: Merck, Darmstadt) are added.
• the sol is stirred for 1 minute at 500 rpm and then treated with ultrasound for 5 minutes.
• the sol-pigment mixture is stirred with a dissolver at approx. 500 rpm until the entire amount has been used up.
• Spraying takes place in a spray tower from Nubilosa, Constance, with a diameter of 0.75 m, a height of 2.5 m and an evaporation rate (based on water) of 1 - 3 liters / hour.
• the air inlet temperature is 270 ° C
• the outlet temperature is approximately 130 ° C.
• the air flow is 7.2 mVmin.
• a two-component nozzle with a spray pressure of 2 bar is used for spraying.
• the delivery rate of the ball valve diaphragm pump used is 60 g sol / min.
• the spray product is captured in a cyclone, precompressed in air at 250 ° C. for 1 hour and then brought to a temperature of 675 ° C.
• a zinc-containing sol is prepared in the same way as in Example 1.
• the following starting materials are weighed in and treated analogously:
• Example 5 The pigment-containing sol from Example 3 is processed analogously to Example 2. However, the compression temperature is 750 ° C.
• Example 5 The pigment-containing sol from Example 3 is processed analogously to Example 2. However, the compression temperature is 750 ° C. Example 5
• the pigment-containing sol from Example 3 is processed analogously to Example 2. However, the compression temperature is 750 ° C and the temperature in the treatment in oxygen is 200 ° C.
• 500 ⁇ l of negative plasma with 10 9 copies of 32 P labeled lambda amplicons are placed in a 2 ml Eppendorf tube.
• 480 ⁇ l binding buffer / proteinase K solution (5: 1) are pipetted in, vortexed and incubated at 70 ° C. for 10 minutes.
• 400 ⁇ l of isopropanolic MGP suspension with a total content of 3 mg MGP are pipetted in.
• mixing is done by vortexing.
• the sample is then placed on a mixer, e.g. B. The ⁇ nomischer 5436 from Eppendorf, incubated.
• the MGP are concentrated by transferring the sample to a magnetic separator. After 1 minute, the supernatant is pipetted off completely.
• 0.5 ml wash buffer is pipetted into the MGPs.
• the sample is vortexed and then transferred to the magnetic separator.
• the supernatant is pipetted off after 1 minute.
• the washing procedure is repeated 2 more times.
• 200 ml of elution buffer are added to the MGP. Incubate at 80 ° C for 10 minutes on a thermal mixer at 1400 RPM.
• the sample is transferred to the magnetic separator and after 1 minute the entire eluate is removed.
• the eluate is then transferred to a new vessel and measured in a scintillation counter.
• the yield can be determined from the ratio of the radioactivity of the eluate to the radioactivity of the sample before the cleaning procedure.
• Example 1 a batch is produced in which the pigment is black mica (BM).
• MMB Microna Matte Black
• Example 9 a batch is produced in which the pigment is MMB (Microna Matte Black (manufacturer: Merck, Darmstadt)).
• MGP according to Examples 7 and 8 is used in sample preparation.
• Human plasma with 100 copies / ml of HCV viruses is used as a sample.
• the eluate from the sample preparation is subjected to ampiification and the amplification result is detected using an electroluminescence method.
• the sample was human plasma with 600 copies ml of HBV viruses.
• a zinc-containing sol is produced in analogy to Example 3, but only 240 g of sol.
• the powder obtained is heated in air at 150 ° C.
• the heating rate is 1 K / min, the holding time is 1 hour.
• the air in the oven is then replaced by N 2 , flushed several times and heated to 700 ° C. with 1 K min, held for 1 hour, cooled to 200 ° C. with 1 K min.
• N 2 is replaced by air and held for 1 hour. Then it is cooled to room temperature. Any aggregates that have arisen are screened with a 50 ⁇ m sieve. The yield is 62.4 g. Screen losses are negligible: aggregates do not occur.
• Example 6 32 P-labeled HIVgag standard with 1.4 Kb is bound to the particles according to Example 10.
• the radioactive measurement shows a binding> 80%.
• the particles in vessel 1 are drawn to the vessel wall with a magnet and then resuspended by shaking. At the same time, the particles are shaken in vessel 2.

Landscapes

• Chemical & Material Sciences (AREA)
• Engineering & Computer Science (AREA)
• Health & Medical Sciences (AREA)
• Life Sciences & Earth Sciences (AREA)
• Genetics & Genomics (AREA)
• Organic Chemistry (AREA)
• Zoology (AREA)
• Biomedical Technology (AREA)
• Wood Science & Technology (AREA)
• Bioinformatics & Cheminformatics (AREA)
• Biotechnology (AREA)
• General Engineering & Computer Science (AREA)
• General Health & Medical Sciences (AREA)
• Analytical Chemistry (AREA)
• Crystallography & Structural Chemistry (AREA)
• Molecular Biology (AREA)
• Biophysics (AREA)
• Microbiology (AREA)
• Physics & Mathematics (AREA)
• Biochemistry (AREA)
• Proteomics, Peptides & Aminoacids (AREA)
• Nanotechnology (AREA)
• Power Engineering (AREA)
• Plant Pathology (AREA)
• Dermatology (AREA)
• Chemical Kinetics & Catalysis (AREA)
• Immunology (AREA)
• Glass Compositions (AREA)
• Glanulating (AREA)
• Treatment Of Liquids With Adsorbents In General (AREA)
• Soft Magnetic Materials (AREA)
• Glass Melting And Manufacturing (AREA)
• Saccharide Compounds (AREA)
• Solid-Sorbent Or Filter-Aiding Compositions (AREA)
• Powder Metallurgy (AREA)
• Apparatus Associated With Microorganisms And Enzymes (AREA)
• Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
• Cleaning And De-Greasing Of Metallic Materials By Chemical Methods (AREA)
• Cleaning Or Drying Semiconductors (AREA)
• Medicines Containing Material From Animals Or Micro-Organisms (AREA)

Priority Applications (12)

Applications Claiming Priority (4)

Related Child Applications (3)

Publications (1)

Family

ID=26050440

Family Applications (1)

Country Status (12)

Cited By (11)

Families Citing this family (23)

Citations (3)

Family Cites Families (3)

• 1999
  • 1999-11-23 DK DK99959300T patent/DK1144620T3/da active
  • 1999-11-23 EP EP99959300A patent/EP1144620B1/de not_active Expired - Lifetime
  • 1999-11-23 WO PCT/EP1999/008996 patent/WO2000032762A1/de active IP Right Grant
  • 1999-11-23 CA CA002352490A patent/CA2352490C/en not_active Expired - Lifetime
  • 1999-11-23 AT AT99959300T patent/ATE248911T1/de active
  • 1999-11-23 AU AU16528/00A patent/AU762188B2/en not_active Expired
  • 1999-11-23 US US09/856,737 patent/US6545143B1/en not_active Expired - Lifetime
  • 1999-11-23 PT PT99959300T patent/PT1144620E/pt unknown
  • 1999-11-23 ES ES99959300T patent/ES2205914T3/es not_active Expired - Lifetime
  • 1999-11-23 DE DE59906909T patent/DE59906909D1/de not_active Expired - Lifetime
  • 1999-11-23 JP JP2000585393A patent/JP3561235B2/ja not_active Expired - Lifetime
• 2001
  • 2001-05-29 NO NO20012620A patent/NO329549B1/no not_active IP Right Cessation
• 2003
  • 2003-02-20 US US10/371,375 patent/US6919444B2/en not_active Expired - Lifetime
  • 2003-12-25 JP JP2003429949A patent/JP3561270B2/ja not_active Expired - Lifetime

Patent Citations (3)

Also Published As

Similar Documents

Legal Events